Foliage blocking agent for precise regulation of expression of cadmium absorption and transportation-related gene in rice and application thereof

ABSTRACT

The present invention relates to the field of environmental protection, and in particular to a leaf surface barrier for ac- curately controlling cadmium absorption and transport related gene expression in rice, and an application thereof. The present inven- tion comprises: reducing a raw material, i.e., selenious acid or selenite, by using ascorbic acid to generate a nanogel, and then emul- sifying the managed for peptization to obtain a leaf surface barrier for accurately controlling cadmium absorption and transport re- lated gene expression in rice; and then mixing the leaf surface barrier with a silica sol to obtain a composite selenium and silica sol leaf surface barrier. The present invention also provides a leaf surface barricading method for accurately controlling cadmium ab- sorption and transport related gene expression in rice. By applying the leaf surface barrier, the silica sol or the composite selenium and silica sol leaf surface barrier having a particular atomization degree and a particular contration by means of foliar spray dur- ing a particular growth period of rice and within a particular time, the cadmium accumulation in rice can be effectively reduced. Ap- plying the present invention to a moderately or lightly polluted field can enable cadmium content in rice to satisfy the food hygiene standards, and therefore is significant for achieving the safe use of the large area of field polluted by heavy metal cadmium in China.

TECHNICAL FIELD

The present invention relates to the field of environmental protection,particularly to a foliage blocking agent for precise regulation of theexpression of the cadmium absorption and transportation-related gene inrice and application thereof.

BACKGROUND ART

As disclosed in “Report on the national general survey of soilcontamination” issued in 2014 by Ministry of Environmental Protection ofthe People's Republic of China and Ministry of Land and Resources of thePeople's Republic of China, the cadmium contaminated soil area accountsfor 7.0% in China, therefore the treatment of the cadmium contaminationin soil is a serious environmental problem to be solved in China,wherein the cadmium contamination is particularly prominent in ricefield.

The treatment technologies against the cadmium contamination in ricefield, are roughly divided into two classes on the basis of thetreatment objectives, a remediation technology of removal of the heavymetals in soil, which has an objective for making the soil quality meetthe environmental standards, and a contamination control technology,which has an objective for guaranteeing the safety of agriculturalproducts.

At present, it has been confirmed that the cadmium hyper-accumulatedplants mainly comprise Sedum alfredii Hance, sedum plumbizincicola,Solanum nigrum L., Thlaspi arvense L., and the like, and the cadmiumenriched plants comprise Amaranthus hypochondriacus L, and the like. Theplant remediation technology requires to occupy farmland and farmingtime, requires a large investment of funds, therefore it is hard to beapplied in a large area. At present, the plant remediation technology isonly suitable for dry lands, and there are no reports on application ofthe same in the flooded rice field yet. In addition, there are noone-to-one correlation between the heavy metal contents in soil and theheavy metal contents in agricultural products, therefore reduction ofthe heavy metal contents in soil to meet the standard level can notguarantee that the safeties of the agricultural products meet thestandard levels.

Obviously, the remediation of the cadmium contamination in rice field isdifficult and presents many technical bottlenecks, therefore the removaltechnique is difficult to solve the cadmium contamination in theregional rice field. It is a practical and feasible technical approachfor improving the quality of the agricultural products by reducing thebioavailability of cadmium, preventing the absorption and accumulationof cadmium in rice, particularly inhibiting the transportation ofcadmium from the rice leaves to the grains.

A new technical idea for controlling the heavy metal contamination infarmlands is disclosed in Chinese Patent Nos. 200610036994.8,201010156359.X, 201010156358.5, 201310737996.X, etc.: the compositesilica sol is directly sprayed on the surface of the rice leaves,without treating the soil, so as to effectively reduce the heavy metalcontents in the agricultural products.

In technical principles, cadmium is a non-essential element in rice,which can compete for the transport proteins for the essential elementsor the beneficial elements, such as Si, Zn, Fe, Mn and the like, so asto enter the roots and be transported upward into the grains (Uraguchi &Fujiwara, 2013). The accumulation of cadmium in rice grains mainlyinvolves in four processes: absorption in the root, transportation inthe xylem, transportation across the vascular bundles, and migrationfrom the phloem to the grains (Uraguchi & Fujiwara, 2012). At present,some important genes which are related to the cadmium absorption in riceand their expression products, the transport proteins, have beenidentified. In the process of the cadmium absorption in the rice roots,the iron transport protein produced by the expression of theconstitutive gene OsNramp5 can absorb Mn and Fe, and simultaneouslyefficiently transport cadmium (Sasaki et al., 2012), the zinc irontransport protein produced by the expression of the genes OsZIP1 andOsZIP3 can transport Zn and Fe, and simultaneously transport cadmium(Ramesh et al., 2003), in addition, the iron transport protein producedby the expression of the Fe-deficiency induction gene OsNramp1, OsIRT1and OsIRT2 can promote the Fe absorption and simultaneously enhance thecadmium absorption (Takahashi et al., 2011; Nakanishi et al., 2006; Lee& An, 2009). In the process of loading in the xylem and upwardtransportation, the transport protein produced by the expression of geneOsHMA2 can load Zn in the xylem, and simultaneously can load cadmium(Nocito et al, 2011; Takahashi et al., 2012), the transport proteinproduced by the expression of genes OsHMA3, OsMTP1, OsABCG43/OsPDR5,etc., can transport the elements, such as zinc, cadmium, and the like,into the vacuoles so as to maintain the equilibrium state of the variouselements in the cells, and inhibit the transportation thereof into thexylem (Ueno et al., 2010; Miyadate et al., 2011; Menguer et al., 2013;Oda et al., 2011). In the process of the loading in the phloem and thetransportation into the grains, mainly the transport proteins such asOsLCT1 and the like, can regulate the transportation of the heavy metalelements, such as manganese, cadmium, etc., from the phloem into thegrains (Uraguchi et al., 2011).

On the basis of the above analysis, the absorption and transportation ofthe non-essential elements, such as cadmium and the like, in the ricebody, can typically be achieved by means of the transport proteins forthe essential elements having similar structures, such as Fe, Zn, Si,and the like, and at present, the specific transport protein which cansolely transport the heavy metal cadmium has not be found yet.Therefore, on the basis of the characteristics that the heavy metalelements compete for the same transport proteins with the essentialelements, the application of the exogenous essential elements canprevent the absorption and in vivo transportation of the heavy metal.

In addition, rice is a typical silicon-philic plant, however, up to now,it is uncertain that silicon is the essential element for plant growth,but there are sufficient evidences showing that silicon is necessary forthe rice production in stable and high yield. Silicon is present in thejuice in the xylem of the plant, mainly in monosilicic acid state[Si(OH)₄], with a small proportion of ionic state Si(H₃SiO₄ ⁻). Theionic state of silicon has a slight high proportion in the rice roots,up to 3%˜8%, dominantly in Si(OH)₄ form; the vast majority of thesilicon in the rice body is present in the form of hydrated amorphoussilica SiO₂.H₂O or polysilicic acid, accounts for about 90%˜95% of thetotal silicon content, and a minority thereof is silicic acid, colloidalsilicic acid and silicate ions.

The absorption of silicon in rice is a typical active absorptionprocess, and silicon is absorbed in the form of silicic acid in the riceroots, therefore such active transportation makes the silicic acidcontent in the solution in the rice xylem is many times higher than thatin the solution outside the roots. At present, three Si transport genes(OsLsi1, OsLsi2 and OsLsi6) have been successfully cloned from the ricesilicon absorption-deficient mutants, wherein the genes OsLsi1 andOsLsi2 are located on chromosomes 2 and 3 respectively, mainly expressedin the roots; the gene OsLsi6 is located on chromosome 6, mainlyexpressed in the leaf sheaths and blades, and sparsely expressed in theroots (Ma et al., 2006; 2007; Yamaji et al., 200). The processe of theabsorption and transportation of silicon in rice involves in 4 steps: inthe first step, the silicic acid can be transported from the outersolution into the cells by the transport protein OsLsi1 in theexodermis, and the silicic acid is released into the apoplast of theaerenchyma by the transport protein OsLsi2; in the second step, thesilicic acid is transported from the apoplast solution into theendodermis cells by the transport protein OsLsi1 in the endodermis, andthe silicic acid is transported into the pericycles by the transportprotein OsLsi2; in the third step, the silicon is transported from thepericycles to the above-ground part through the xylem vessel, in thenon-polymeric monosilicic acid form, with the transpiration stream; inthe fourth step, the transport protein OsLsi6 which is positioned in thexylem parenchyma cells close to the side of the vessels in the leafsheaths and blades is responsible for unloading and distributing thesilicon in the xylem, and dehydrated and polymerized under thetranspiration effect, so as to form silica gel (SiO₂.nH₂O), which isdeposited in the cell wall and the intercellular space in the differenttissues and organs in the above-ground part, wherein more than 90% ofsilicon is present, in silica gel form, in rice.

Silicon is an important constituent element in rice body, and also amain constituent component in the cell wall of rice. After deposited inthe apoplast (the cell wall and the intercellular space), silicon cannot only reduce the cell voidage, but also can co-precipitate withcadmium and bind the same within the cell wall, so as to reduce thetransportation of cadmium from the apoplast into the cell (Shi et al.2005; Shi et al., 2010). Silicon can also improve the oxidative stressresistance in rice, alleviate the cadmium stress toxicity, and promotethe increase of biomass (Liang et al., 2007), so that the exogenousapplication of silicon can improve the biomass of rice, andsimultaneously reduce the cadmium content in the grains.

On the basis of the above mechanism of silicon, it is clear that siliconcan prevent the accumulation of cadmium in the grains, so that siliconis determined to be used as a main exogenous regulator for preventingcadmium in rice grains. The above four authorized patents are thetechnologies for preventing the accumulation of cadmium in rice grainsusing the silica sol as the exogenous regulator, and provide theprocesses for preparing the silica sol, the rare-earth composite silicasol, the selenium dioxide composite silica sol.

However, the above four patents do not involve in how the silicon andselenium affect the active expression of the cadmium absorption andtransportation-related gene in rice, how the foliage blocking agents areapplied to achieve the precise regulation of the active expression ofthe cadmium absorption and transportation-related genes in rice, so asto inhibit the accumulation of cadmium in rice. These patents have notprovided the precise regulation technologies and methods on theseaspects yet.

CONTENTS OF THE INVENTION

In order to overcome the disadvantages and deficiencies in the priorart, the primary object of the present invention is to provide a methodfor preparing a foliage blocking agent for precise regulation of theexpression of the cadmium absorption and transportation-related gene inrice.

Another object of the present invention is to provide a foliage blockingagent prepared by the above method.

Yet another object of the present invention is to provide a seleniumcomposite silica sol foliage blocking agent for precise regulation ofthe expression of the cadmium absorption and transportation-related genein rice.

The fourth object of the present invention is to provide the applicationof the above foliage blocking agent and/or selenium composite silica solfoliage blocking agent.

The fifth object of the present invention is to provide a foliageblocking method for precise regulation of the expression of the cadmiumabsorption and transportation-related gene in rice, which can preciselyregulate the transportation of cadmium from the rice blades into thegrains.

The objects of the present invention are achieved by the followingtechnical solutions.

A method for preparing a foliage blocking agent for precise regulationof the expression of the cadmium absorption and transportation-relatedgene in rice, comprises the steps of

(1) preparation of nano-sized selenium gel: an ascorbic acid solution isadded to a selenium-containing solution in a water bath at 25˜50° C.,reacted for 2˜5 h, separated the products, and purified, so as to obtaina nano-sized selenium gel;

(2) dispergation of the nano-sized selenium gel: an emulsifying agent isadded to the nano-sized selenium gel prepared in step (1) understirring, adjusted the pH to 4.5˜6.5; so as to obtain a foliage blockingagent for precise regulation of the expression of the cadmium absorptionand transportation-related gene in rice;

wherein, the selenium-containing compound in the selenium-containingsolution in step (1) is preferably at least one from selenious acid,sodium selenite and potassium selenite;

the selenium content in the selenium-containing solution in step (1) ispreferably 0.1˜5% by mass, more preferably 0.5˜1.5% by mass;

the ascorbic acid content in the ascorbic acid solution in step (1) ispreferably 0.5˜10% by mass, more preferably 1˜3% by mass;

the molar ratio between the ascorbic acid in the ascorbic acid solutionand the selenium in the selenium-containing solution in step (1) is(1:1)˜(1:3);

the separation of the product in step (1) is preferably carried out bycentrifugation;

the centrifugation is preferably carried out at a centrifugal force of4000˜10000 g, for 10˜40 min;

the purification in step (1) is preferably carried out by washing theproduct with water and precipitating 2-5 times;

the final emulsifying agent content in step (2) is preferably 0.05˜5% bymass;

the emulsifying agent in step (2) is preferably at least one frompolyvinylpyrrolidone, polyvinyl alcohol, Tween series, and Span series;

the selenium content in the foliage blocking agent in step (2) is0.15˜2% by mass.

A foliage blocking agent for precise regulation of the expression of thecadmium absorption and transportation-related gene in rice is preparedby the above method.

A selenium composite silica sol foliage blocking agent for preciseregulation of the expression of the cadmium absorption andtransportation-related gene in rice, comprises the above foliageblocking agent and a silica sol.

A method for preparing the selenium composite silica sol foliageblocking agent for precise regulation of the expression of the cadmiumabsorption and transportation-related gene in rice, comprises the stepsof:

the above foliage blocking agent and a silica sol are mixed to uniform,so as to obtain a selenium composite silica sol foliage blocking agentfor precise regulation of the expression of the cadmium absorption andtransportation-related gene in rice;

the foliage blocking agent and the silica sol have a volume ratio of(1:1)˜(1:4);

the silica content in the selenium composite silica sol foliage blockingagent is 3˜10% by mass;

the mass ratio between the selenium and the silica in the seleniumcomposite silica sol foliage blocking agent is (1:10)˜(1:55);

the silica sol is preferably an acidic silica sol;

the acidic silica sol has a pH of 4.5˜6.5, and a silica content of15%˜20% by mass.

A method for preparing the acidic silica sol comprises the steps of:

a metallic silicon powder or a metal silicate are slowly added to analkaline solution in a water bath at 40˜60° C., under a stirring speedof 0.5˜5 m/s, so as to obtain an alkaline silicon-containing solution;the alkaline silicon-containing solution is passed through ahydrogen-type weak acid cation exchange resin column at a speed of 1˜10mL/min, controlled the pH of the collection liquid at the column outletto 4.5˜6.5, so as to obtain an active acidic silica sol precursor; theactive acidic silica sol precursor is heated to 40˜60° C. in amicrowaver or water bath, and maintained the temperature, stirred for10˜30 min, then stood, cooled, and aged for 30˜60 min, so as to preparethe acidic silica sol;

the metal silicate is Na₂SiO₃, K₂SiO₃ or Li₂SiO₃, preferably Na₂SiO₃;

the alkaline silicon-containing solution has a concentration of 5% bymass to saturation, and a pH of 10˜13;

the metallic silicon powder or the metal silicate requires a lowimpurity content;

the alkaline solution is sodium hydroxide solution, potassium hydroxidesolution or aqueous ammonia, preferably aqueous ammonia;

the alkaline solution has a concentration of 0.1˜5 M, preferably 0.2˜2M.

A selenium composite silica sol foliage blocking agent is prepared bythe above method.

The application of the foliage blocking agent and/or the seleniumcomposite silica sol foliage blocking agent in the technical filed forpreventing heavy metal in plant.

A foliage blocking method for precise regulation of the expression ofthe cadmium absorption and transportation-related gene in rice,comprises the steps of:

the above foliage blocking agent, silica sol or selenium compositesilica sol foliage blocking agent are diluted with water, in the periodfrom the rice tillering stage to the jointing stage or from the headingstage to the filling stage, atomized and sprayed on the rice leaf bladesonce, or sprayed once for each period; the spraying is carried out in asunny day or at 4˜6 p.m. in a cloudy day;

the selenium content in the diluted foliage blocking agent is controlledat 0.01% to 0.1% by mass, preferably 0.05% by mass;

the silica content in the diluted silica sol or selenium compositesilica sol foliage blocking agent is controlled at 0.1% to 0.1% by mass,preferably 0.5% by mass;

the spraying dosage in the period from the tillering stage to thejointing stage is 80˜120 liters of the diluted foliage blocking agent,silica sol, or selenium composite silica sol foliage blocking agent permu;

the spraying dosage in the period from the heading stage to the fillingstage is 100˜150 liters of the diluted foliage blocking agent, silicasol, or selenium composite silica sol foliage blocking agent per mu;

the atomization and spraying are preferably carried out as follows: theatomized droplets are controlled to have a particle size of less than1000 microns, uniformly sprayed on both the front and the back surfacesof the rice blade.

The principles of the invention are as follows:

On the basis of the transport mechanism of the beneficial elements, suchas silicon, selenium, and the like, in rice, and the transport mechanismof the heavy metal cadmium in rice, starting from these aspects, such asregulation of the distribution between the stems and the leaves,regulation of the activity of the transport protein, competition for thetransportation channel, and the like, the key genes for controlling thetransportation of cadmium from the rice blade into the grains,comprising OsLsi1, OsLsi2, OsHMA3, OsNramp5 and OsLCT1, are analyzed.The silica sol, selenium sol and selenium composite silica sol foliageblocking agents are developed, all of them can efficiently and preciselyregulate the activity of the above key genes; as compared with those ofspraying clean water, spraying the above sol materials on the leafblades can efficiently reduce the cadmium content in rice. Secondly, theexpression data analysis on the cadmium/silicon/selenium-relatedtransport proteins queried and extracted from GenBank and RiceXProdatabases show that the key step for the accumulation of cadmium in therice grains lies in transportation of cadmium from the phloem to thegrains. At present, it is found that the transport protein in the phloemrelated to the accumulation of cadmium in the rice grains is mainlyOsLCT1. On the basis of the expression pattern analysis ofbioinformatics, it is found that the gene OSLCT1 in the Oryza. Sativa L.spp. japonica is mainly expressed on the leaf blades, sparsely expressedin roots; in addition, the gene OSLCT1 not only shows differentexpressions in the different growth periods, wherein, during thetransformation from the vegetative growth phase to the reproductivegrowth stage (tillering stage-heading stage), the expression level issignificantly increased, and reached the maximum in the maturationstage; but also show a certain periodicity in its expression per day,wherein the expression level is significantly increased at 4˜6 p.m. perday, and reached the maximum at 8˜10 p.m., and the expression level islow in the rest period. The optimal technology for applying the aboveblocking agent materials is determined on the basis of the temporal andspatial specific expression patterns and the temporal and spatialexpression specificity of various transport proteins in above analysis,so as to achieve the precise prevention of the transportation of cadmiumfrom the rice blade to the grains.

As compared with the prior art, the present invention has the advantagesand effects as follows:

(1) The present invention provides a foliage blocking method for preciseregulation of the expression of the cadmium absorption andtransportation-related gene in rice, wherein the activity of the cadmiumabsorption and transportation-related gene in rice is preciselyregulated to inhibit the accumulation of cadmium in rice. As thephysiological mechanism of high efficient blocking of the cadmiumaccumulation in rice by spraying the materials, such as selenium,silicon and the like on the leaf blades can be explained from thetechnical principles, and the field application of the blockingmaterials can be taught from the present invention, the foliage blockingmethod determined by the temporal and spatial expression data analysison the cadmium/silicon/selenium-related transport genes in the presentinvention can efficiently inhibit the accumulation of cadmium in rice.

(2) The object of the present invention is to make the safety of thecadmium content in rice and the safety of the cadmium contaminated ricefield meet the standards. It is hard to remove the selenium in soil inthe large area of the cadmium contaminated rice field by using theprocessing methods, such as physical removal, chemical leaching, and thelike. Starting from the basic theories, such as the mechanisms oftransportation of the beneficial elements, such as silicon, selenium andthe like, and the heavy metal cadmium, in rice, the present inventionprovides applicable key products, such as foliage blocking agent(selenium sol) and selenium composite silica sol foliage blocking agentas well as application methods thereof, in order to reduce thetransportation of cadmium from the rice blades to the grains, thusincreasing the safety of the rice. This control technology is one thatis suitable for the low-to-moderate cadmium contaminated rice fields inChina, which can provide technical supports for treating the large areaof the cadmium contaminated rice fields in China.

(3) The present invention provides a selenium sol and a seleniumcomposite silica sol foliage blocking agent, as well as a foliageblocking method, which has the characteristics, such as low cost andhigh efficiency.

(4) The present invention provides the products, such as silica sol,selenium sol foliage blocking agent, and selenium composite silica solfoliage blocking agent, the cost for spraying these blocking agents onthe leaf blades is only 500˜1500 yuan/hectare, which is 10% of that ofapplying the soil passivating agent, and has an equivalent or evenbetter effect in reducing the cadmium content in rice.

DESCRIPTION OF FIGURES

FIG. 1 is a transmission electron microscopic image of an acidic silicasol.

FIG. 2 is a diagram showing the particle size distribution of thefoliage blocking agent (selenium sol).

FIG. 3 is a transmission electron microscopic image of the foliageblocking agent (selenium sol).

FIG. 4 is a transmission electron microscopic image of the seleniumcomposite silica sol foliage blocking agent.

FIG. 5 is a diagram showing the particle size distribution of theselenium composite silica sol foliage blocking agent.

PARTICULAR EMBODIMENTS

The present invention will be further described in details below incombination with the examples and accompanying drawings, but theembodiments of the present invention are not limited thereto.

Method for culturing the suspension cell of rice (Oryza sativa L.Japonica) comprises the steps of: the matured and plump seeds of Oryzasativa L. Japonica were selected, and sterilized with 70% by volume ofalcohol or 30% by mass of sodium hypochlorite; uniformly spread theseeds on the plates containing a solid culture medium with forceps,cultured in an incubator, in dark, at a culture temperature of 25□, fora period of time, then taken out the hypocotyls with forceps, culturedon another solid culture medium, so as to obtain callus tissues. 10 g ofthe callus tissues were broken, added to the conical flasks containing aliquid culture medium (50 mL), cultured in a shaker at a temperature of28□, for about 1 month, so as to obtain the suspension cells of therice.

EXAMPLE 1 Preparation of Acidic Silica Sol

(1). 200 mL water was metered, and added 0.8 g of sodium hydroxide, soas to formulate 0.1 M sodium hydroxide solution; actuated a stirrer upto a speed of 0.5 m/s, and heated to 40° C., then slowly added 116 gNa₂SiO₃, fully dissolved, then cooled to the room temperature, so as toformulate an alkaline silicon solution with a pH of 10; passed thealkaline silicon solution through 100 mL (wet volume) hydrogen-type weakacid cation resin exchange column at an uniform speed of 1 mL/min, andcontrolled the pH value of the collection liquid at the column outlet to4.5, so as to obtain an active acidic silica sol precursor; stirred theactive acidic silica sol precursor in a water bath and heated to 40° C.,maintained the temperature and stirred for 30 minutes, stood, cooled andaged for 60 min, so as to obtain an acidic silica sol, for use; theacidic silica sol having a pH of 4.5, and a silica content of 18% bymass;

(2) 200 mL water was metered, added 56 g potassium hydroxide, so as toformulate 5 M potassium hydroxide solution; actuated a stirrer up to aspeed of 5 m/s, and heated to 60° C., then slowly added 110 g Li₂SiO₃,fully dissolved, then cooled to the room temperature, so as to preparean alkaline silica solution with a pH of 12.5; passed the alkalinesilica solution through 100 mL (wet volume) hydrogen-type weak acidcation resin exchange column at an uniform speed of 10 mL/min, andcontrolled the pH of the collection liquid at the column outlet to 6.5,so as to obtain an active acidic silica sol precursor; stirred theactive acidic silica sol precursor in a water bath and heated to 60□,maintained the temperature and stirred for 10 min, stood, cooled andaged for 30 min, so as to obtain an acidic silica sol, for use; theacidic silica sol having a pH of 6.5, and a silica content of 20% bymass;

(3) 200 mL water was metered, added 7 g aqueous ammonia, so as toformulate 1M ammonium hydroxide solution; actuated a stirrer up to aspeed of 2 m/s and heated to 45□, then slowly added 90 g Na₂SiO₃, fullydissolved, then cooled to the room temperature, so as to prepare analkaline silica solution with a pH of 11; passed the alkaline silicasolution through 100 mL (wet volumn) hydrogen-type weak acid cationresin exchange column at an uniform speed of 5 mL/min, controlled the pHvalue of the collection liquid at the column outlet to 5.5, so as toobtain an active acidic silica sol precursor; stirred the active acidicsilica sol precursor in a water bath and heated to 45□, maintained thetemperature and stirred for 20 min, stood, cooled and aged for 45 min,so as to obtain an acidic silica sol, for use; the acidic silica solhaving a pH of 5.5, and a silica content of 15% by mass, with a TEMimage shown in FIG. 1.

EXAMPLE 2 Preparation of Selenium Sol Foliage Blocking Agent

(1) a selenious acid solution was formulated, and controlled theselenium content at 0.1% by mass;

(2) 0.5% by mass of the ascorbic acid solution was added into theselenious acid solution prepared in step (1) in a water bath at 25° C.,controlled the molar ratio between ascorbic acid and selenium to 1:1,and reacted for 2 h; then centrifuged in a centrifuge at 10000 g, for 10min; followed by washing the precipitates three times with ultra-purewater, so as to obtain a nano-sized selenium gel;

(3) the emulsifying agent, polyvinylpyrrolidone, was added to thenano-sized selenium gel prepared in step (2), under stirring, whereinthe final emulsifying agent content was 0.05% by mass, and adjusted thepH to 4.5, so as to obtain a foliage blocking agent (selenium solfoliage blocking agent) for precise regulation of the expression of thecadmium absorption and transportation-related gene in rice, wherein theselenium sol foliage blocking agent had a selenium content of 0.15% bymass.

EXAMPLE 3 Preparation of Selenium Sol Foliage Blocking Agent

(1) a sodium selenite solution was prepared, and controlled the seleniumcontent at 5% by mass;

(2) 10% by mass of the ascorbic acid solution was added to the sodiumselenite solution prepared in step (1) in a water bath at 50° C.,controlled the molar ratio between ascorbic acid and selenium to 1:3,and reacted for 5 h; then centrifuged in a centrifuge at 4000 g for 40min, followed by washing the precipitates 3 times with ultra-pure water,so as to obtain a nano-sized selenium gel;

(3) an emulsifying agent, Span-80, was added to the nano-sized seleniumgel prepared in step (2), under stirring, wherein the final emulsifyingagent content was 5% by mass, and adjusted the pH to 6.5, so as toobtain a foliage blocking agent (selenium sol foliage blocking agent)for precise regulation of the expression of the cadmium absorption andtransportation-related gene in rice, wherein the selenium sol foliageblocking agent had a selenium content of 2% by mass.

EXAMPLE 4 Preparation of Selenium Sol Foliage Blocking Agent

(1) a potassium selenite solution was formulated, and controlled theselenium content at 1.5% by mass;

(2) 2.5% by mass of the ascorbic acid solution was added to thepotassium selenite solution prepared in step (1) in a water bath at 50°C., controlled the molar ratio between ascorbic acid and selenium to1:1.5, and reacted for 3 h; then centrifuged in a centrifuge at 5000 gfor 30 min, followed by washing the precipitates 3 times with ultra-purewater, so as to obtain a nano-sized selenium gel;

(3) an emulsifying agent, Tween-80, was added to the nano-sized seleniumgel prepared in step (2), under stirring, wherein the final emulsifyingagent content was 1.5% by mass, and adjusted the pH to 5.5, so as toobtain a foliage blocking agent (selenium sol foliage blocking agent)for precise regulation of the expression of the cadmium absorption andtransportation-related gene in rice, wherein the selenium sol foliageblocking agent had a selenium content of 1.5% by mass.

(4) the selenium sol foliage blocking agent samples prepared in step (3)were carried out a particle size distribution analysis, the resultsshowed that the average particle diameter was 17.35±1.4 nm, as shownFIG. 2; the prepared selenium sol foliage blocking agent samples werecarried out a morphology analysis by TEM, the nano-sized selenium formedin the reaction system was a spherical particle, and uniformlydispersed, having an average particle diameter of about 15 nm, as shownin FIG. 3.

EXAMPLE 5 Preparation of Selenium Composite Silica Sol Foliage BlockingAgent

The selenium sol foliage blocking agent prepared in example 4 was mixedwith the acidic silica sol, having a pH of 5.5, and silica content of15% by mass, the acidic silica sol, having a pH of 4.5, and silicacontent of 18% by mass, and the acidic silica sol, having a pH of 6.5,and silica content of 20% by mass, prepared in example 1, respectively,wherein the volume ratio between the selenium sol foliage blocking agentand the silica sol was 1:1, 1:2 and 1:4 respectively, stirred for 30min, so as to obtain a selenium composite silica sol foliage blockingagent for precise regulation of the expression of the cadmium absorptionand transportation-related gene in rice; the silica content in theprepared selenium composite silica sol foliage blocking agent was 7.5%,6% and 4% by mass respectively, and the mass ratio between the seleniumand silica were 1:10, 1:24 and 1:53 respectively. The prepared seleniumcomposite silica sol foliage blocking agent samples having a silicacontent of 7.5% by mass were carried out a morphology analysis by TEM,as shown in FIG. 4, the nano-sized selenium formed in the reactionsystem was loaded on the surface of the nano-sized silica sphere, havinga particle size of 45˜55 nm (FIG. 5).

EXAMPLE 6 Precise Regulation of the Acidic Silica Sol on the Expressionof the Cadmium Transport Gene in Rice Suspension Cell and Inhibition ofthe Same on the Cadmium Absorption

The silica sol prepared in example 1 having a pH of 5.5 and a silicacontent of 15% by mass, was diluted to a silica content of 0.5% by mass,and used to treat the rice cells (Oryza sativa L. Japonica) for 24 h,then added various concentrations of cadmium, and continued to culturefor 24 h, then extracted the RNA of the rice suspension cells, andcarried out qRT-PCR analysis (the corresponding primers shown in table14). The results showed that as compared with the blank control (CK,without addition of silica sol), at the same cadmium concentration,after treating with the silica sol, the expression levels of the genesOsLsi1, OsNramp5 and OsLCT1 in the rice cells were significantly reduced(table 1, table 2, table 3), and the expression level of the gene OsHMA3was significantly increased (table 4). The proteins expressed by thegenes OsLsi1 and OsNramp5 may be associated with the absorption ofcadmium, the proteins expressed by the gene OsLsi1 may be associatedwith the transportation of cadmium from the phloem to the grains, thedown-regulations of the expression of the three genes facilitated thereduction of the absorption of cadmium in rice and the reduction of thetransportation of cadmium into the grains; and the protein expressed bygene OsHMA3 may have the functionality which was associated with thetransportation of cadmium into the vacuoles, so as to reduce thetransportation of the same into the xylem, and in turn reduce thetransportation of cadmium from the roots to the above-ground part. Afterthe silica sol was used to precisely regulate the above genes, thecadmium accumulation levels in the rice cells were significantlyreduced, as compared with the controls, after treating with silica solsat 10, 20, 40 μM Cd, the cadmium concentrations in the rice cells werereduced by 72.8%, 75.5% and 81.3% respectively (table 5).

TABLE 1 The effects of the silica sol treatment on the expression levelsof the transport gene OsLsi1 in the rice suspension cells Averagereduction as Silica sol compared with the control Blank (CK) treatment(%) 0  4.13 ± 0.81  3.66 ± 0.75 11.4 Cd (10 μM) 15.62 ± 1.32 13.25 ±0.92 15.2 Cd (20 μM) 19.85 ± 1.25 16.43 ± 0.75 17.2 Cd (40 μM) 22.78 ±0.97 17.65 ± 0.84 22.5

TABLE 2 The effects of the silica sol treatment on the expression levelsof the cadmium transport gene OsNramp5 in the rice suspension cellsAverage reduction as Silica sol compared with the control Blank (CK)treatment (%) 0 2.8 ± 0.25 1.9 ± 0.36 32.1 Cd (10 μM) 4.2 ± 0.32 3.5 ±0.29 16.7 Cd (20 μM) 5.6 ± 0.28 4.1 ± 0.35 26.8 Cd (40 μM) 6.8 ± 0.454.6 ± 0.42 32.4

TABLE 3 The effects of the silica sol treatment on the expression levelsof the cadmium transport gene OsLCT1 in rice suspension cells. Averagereduction as compared with the Blank (CK) Silica sol treatment control(%) 0 1.85 ± 0.09 1.43 ± 0.06 22.7 Cd (10 μM) 1.52 ± 0.04 0.71 ± 0.0753.3 Cd (20 μM) 1.44 ± 0.04 0.58 ± 0.05 59.7 Cd (40 μM) 1.32 ± 0.06 0.67± 0.08 49.2

TABLE 4 The effects of the silica sol treatment on the expression levelsof the cadmium transport gene OsHMA3 in rice suspension cells Averageincrease as Silica sol compared with the control Blank (CK) treatment(%) 0 2.25 ± 0.15 3.11 ± 0.32 38.2 Cd (10 μM) 1.84 ± 0.18 3.07 ± 0.2166.8 Cd (20 μM) 2.66 ± 0.21 3.26 ± 0.3  22.6 Cd (40 μM) 3.05 ± 0.26 3.64± 0.31 19.3

TABLE 5 The effects of the silica sol treatment on the cadmium contentsin the rice cells (μg kg⁻¹) Average reduction as Silica compared withthe control Blank (CK) sol treatment (%) Cd (10 μM)  9.2 ± 0.8 2.5 ± 0.672.8 Cd (20 μM) 18.4 ± 1.1 4.5 ± 0.7 75.5 Cd (40 μM) 36.4 ± 6.8 6.8 ±0.9 81.3

EXAMPLE 7 Precise Regulation of the Selenium Sol Foliage Blocking Agenton the Expression of the Cadmium Transport Gene in the Rice SuspensionCells and Inhibition of the Same on the Cadmium Absorption

The selenium sol foliage blocking agents prepared in example 4 werediluted to various concentrations respectively (0.01%, 0.05%, 0.1%), andcultured the rice suspension cells (Oryza sativa L. Japonica) for 24 h,then added various concentrations of cadmium (10 μM, 20 μM and 40 μM),and cultured for 24 h, extracted the RNA from the rice cells, andcarried out reverse transcription, then detected the expression levelsof the genes OsNramp5 and OsHMA3 (the corresponding primers shown intable 14). The results showed that (table 6, table 7), as compared withthe blank (CK, without addition of selenium sol foliage blocking agent),the expression levels of the gene OsHMA3 in the rice suspension cellswere up-regulated in duplications, and the expression levels of the geneOsNramp5 in the rice suspension cells were significantly suppressed; athigh cadmium concentration (40 μM), after treating with 0.05% seleniumsol foliage blocking agent, the suppression effects on the expression ofthe gene OsNramp5 were optimal. The protein expressed by the geneOsNramp5 may be associated with the absorption of cadmium, and theprotein expressed by the gene OsHMA3 may have the functionality whichmay be associated with the transportation of cadmium into the vacuoles,so as to reduce the transportation of the same into the xylem, and inturn reduce the transportation of cadmium from the roots to theabove-ground part. After the selenium sol foliage blocking agent wasused to precisely regulate the above genes, the cadmium accumulationlevels in the rice cells were significantly reduced, and as comparedwith the control, at 10, 20, 40 μM Cd, after treating with 0.05%selenium sol foliage blocking agent, the cadmium concentrations in therice cells were reduced by 47.1%, 55.3% and 61.7% respectively (table8).

TABLE 6 The effects of the selenium sol foliage blocking agent on theexpression levels of the cadmium transport protein OsNramp5 in the ricesuspension cells. Blank Selenium Selenium Selenium (CK) (0.01%) (0.05%)(0.1%) 0 2.11 ± 0.15 1.82 ± 0.04 1.97 ± 0.08 1.76 ± 0.09 Cd (10 μM) 4.23± 0.28 1.96 ± 0.09 1.34 ± 0.06 0.84 ± 0.04 Cd (20 μM) 6.72 ± 0.05 1.55 ±0.08 0.92 ± 0.07 0.63 ± 0.04 Cd (40 μM) 7.96 ± 0.08 1.34 ± 0.05 0.85 ±0.08 1.55 ± 0.06

TABLE 7 The effects of the selenium sol foliage blocking agent on theexpression levels of the cadmium transport protein OsHMA3 in the ricesuspension cells Selenium Selenium Selenium Blank (CK) (0.01%) (0.05%)(0.1%) 0 1.21 ± 0.15 1.56 ± 0.18 1.85 ± 0.18 2.54 ± 0.21 Cd (10 μM) 1.86± 0.32 2.25 ± 0.28 3.14 ± 0.29 3.81 ± 0.26 Cd (20 μM) 2.26 ± 0.25 2.81 ±0.28 3.62 ± 0.28 4.97 ± 0.28 Cd (40 μM) 2.45 ± 0.31 3.26 ± 0.31 3.87 ±0.24 4.52 ± 0.24

TABLE 8 The effects of 0.05% selenium sol foliage blocking agenttreatment on the cadmium contents in the rice cells (μg kg⁻¹). Averagereduction as Selenium compared with the control Blank (CK) (0.05%) (%)Cd (10 μM)  8.7 ± 0.5 4.6 ± 0.4 47.1 Cd (20 μM) 15.2 ± 0.9 6.8 ± 0.755.3 Cd (40 μM) 29.8 ± 3.2 11.4 ± 1.2  61.7

EXAMPLE 8 Precise Regulation of the Selenium Sol Foliage Blocking Agent,Silica Sol, and Selenium Composite Silica Sol Foliage Blocking Agent onthe Expression of the Cadmium Transport Gene in Rice and Inhibition ofthe Same on the Cadmium Absorption Under Pot Culture Conditions

The rice (a variety of YouYou 128) was cultured in Kimura culture mediumfor 3 weeks, then transferred into a culture solution containing 5 μMCd, and carried out the following treatments: (1) treatments withcontrol (CK): 200 mL deionized water was sprayed on each pot; (2)treatments by spraying the silica sol foliage blocking agent on the leafblades: 200 mL the diluted silica sol (the silica sol prepared inexample 1, having a pH of 5.5 and a silica content of 15% by mass, thendiluted to a silica content of 0.5% by mass) was sprayed on each pot;(3) treatments by spraying the selenium sol foliage blocking agent onthe leaf blades: 200 mL the diluted selenium sol foliage blocking agent(prepared in example, then diluted to a selenium content of 0.05%) wassprayed on each pot; (4) treatments by spraying the selenium compositesilica sol foliage blocking agent on the leaf blades: 200 mL theselenium composite silica sol foliage blocking agent was sprayed on eachpot (prepared in example 5, then diluted to a silica content of 0.5% anda selenium content of 0.05%). After one week, the samples werecollected, and tested the relative expression levels of the genes OsLsi1and OsLsi2 in the roots (corresponding primers shown in table 14), andtested the cadmium contents in the stems and leaves. The results showedthat all of the three sol foliage blocking agents can precisely inhibitthe relative expression levels of the genes OsLsi1 and OsLsi2 (table 9),so as to reduce the cadmium contents in the rice stems and leaves. Ascompared with the control, after the treatments by spraying the silicasol, selenium sol and selenium composite silica sol on leaf blades, thecadmium accumulation levels in the above-ground part of rice werereduced by 63.8%, 51.6%, 66.1% respectively; and the selenium compositesilica sol had the optimal effects in inhibiting the cadmiumaccumulation in the above-ground part of rice (table 10).

TABLE 9 The effects of various foliage blocking materials on therelative expression levels of the genes OsLsi1 and OsLsi2 in the riceroots OsLsi1 OsLsi2 Control (CK) Relative expression level    1 ± 0.0282   1 ± 0.1263 Silica sol Relative expression level 0.485 ± 0.118 0.629 ±0.125 Reduction as compared with control (%) 51.45 37.07 Selenium solRelative expression level 0.486 ± 0.103 0.326 ± 0.030 Reduction ascompared with control (%) 51.39 67.4  Selenium composite silica solRelative expression level 0.451 ± 0.078 0.314 ± 0.078 Reduction ascompared with control (%) 54.86 68.53

TABLE 10 The effects of various foliage blocking materials on thecadmium accumulation levels in the rice stems and leaves Reduction asCadmium level compared with control (mg/kg) (%) CK 3.45 ± 0.47 Silicasol 1.25 ± 0.11 63.8 Selenium sol 1.67 ± 0.27 51.6 Selenium compositesilica sol 1.17 ± 0.09 66.1

EXAMPLE 9 The Effects of the Selenium Sol Foliage Blocking Agent, SilicaSol and Selenium Composite Silica Sol Foliage Blocking Agents on theReduction of the Cadmium Accumulation in the Rice Grains in the FieldConditions.

The test time was from April to November, 2014, the test site waslocated in a Cd contaminated rice field in Hongxing village, Dongtangtown, Renhua county, Shaoguan city, Guangdong province, China. Thesurface soils were collected from this plot (0˜30 cm) for analysis. Thesoils had a pH of about 5.86, a Cd content of about 1.85 mg kg⁻¹, and anAs content of 18.20 mg kg⁻¹. This plot was a typical cadmiumcontaminated rice field resulting from mining The rices to be testedwere the early mature variety of WuYou 613 in 2014, and the late maturevariety of WuFengYou 615.

Four treatments were set for the tests as follows: (1) a blank control,(CK), i.e., spraying an equal amount of clean water; (2) spraying thesilica sol foliage blocking agent (prepared in example 1, having a pH of5.5 and a silica content of 15% by mass) on the leaf blades; (3)spraying the selenium sol foliage blocking agent (prepared in example 4)on the leaf blades; (4) spraying the selenium composite silica solfoliage blocking agent (prepared in example 5) on the leaf blades. Eachtreatment was repeated 3 times, arranged in random, tested 12 plots intotal, wherein each plot had an area of 5*64=30 m², and was ensured tobe separately irrigated and drained.

Spraying Technology:

In the early mature rices, the blocking agents were sprayed twice foreach of the treatments, once during the period from the tillering stageto the jointing stage, the other once during the period from the headingstage to the filling stage. During the period from the tillering stageto the jointing stage, the sprayed silica sol had a silica content of0.1%, the selenium sol had a selenium content of 0.01%, and the seleniumcomposite silica sol had a silica content of 0.1% (prepared in example5, having a silica content of 7.5% by mass, a mass ratio between silicaand selenium of 10:1), and during the period from heading stage to thefilling stage, the sprayed silica sol had a silica content of 0.5%, andthe selenium sol had a selenium content of 0.05%, the selenium compositesilica sol had a silica content of 0.5% (prepared in example 5, having asilica content of 7.5% by mass, and a mass ratio between silica andselenium of 10:1). During the period from the tillering stage to thejointing stage, the spraying dosage for each of the blocking agents was100 liters/mu, the spraying was carried out at 4:00 p.m. in a sunny day;during the period from the heading stage to the filling stage, thespraying dosage of each of the blocking agents was 150 liters/mu, thespraying was carried out at 6:00 p.m. in a cloudy day. During spraying,the atomized droplets were controlled to have a particle size of lessthan 1000 microns, and uniformly sprayed on both the front and backsurfaces of the rice blade.

In the late mature rice, the blocking agent was sprayed once for each ofthe treatments. The spraying was carried out during the period from thetillering stage to the jointing stage, and the sprayed silica sol had asilica content of 1%, the selenium sol had a selenium content of 0.1%,and the selenium composite silica sol had a silica content of 1%(prepared in example 5, having a silica content of 7.5% by mass, and amass ratio between silica and selenium of 10:1); the spraying dosage foreach of the blocking materials was 120 liters/mu, the spraying wascarried out at 5:00 p.m. in a sunny day. During spraying, the atomizeddroplets were controlled to have a particle size of less than 1000microns, and uniformly sprayed on both front and back surfaces of therice blade.

As shown in tables 11-12, after the treatments by various of foliageblocking agents, both the early and late mature rices in 2014 had anincreased yield; wherein the early mature rice had much more increaselevel than that of the late mature rice, and after the treatment withthe selenium composite silica sol, both the early and late mature riceshad a significant increase level in rice yield, which was increased by7.32% and 4.85% respectively as compared with the control.

TABLE 11 The statistical results for the yields of the early mature ricein 2014 Plot yield (wet Plot average Dry field Yield yield, kilogram)yield per mu increase □ □ □ (kilogram) (kilogram) (%) control 37 37.2538.5 37.58a 647.2 Silica sol 38 39.25 40.25 39.17ab 674.6 4.23 Seleniumsol 38 40.2 39.5 39.23ab 675.7 4.41 Selenium 40 41 40 40.33b 694.6 7.32composite silica sol

TABLE 12 The statistical results for the yields of the late mature ricein 2014 Plot yield (wet Plot average Dry yield Yield yield, kilogram)yield per mu increase □ □ □ (kilogram) (kilogram) (%) Control 33.5 34.533.5 33.83a 585.5 Silica sol 32.5 37.5 34 34.6a 598.8 2.27 Selenium sol33.7 34.7 35.1 34.5a 597.1 1.98 Selenium 34 38 34.25 35.43b 613.9 4.85composite silica sol

As shown in table 13, the treatments by spraying the foliage blockingagent can significantly reduce the cadmium content in rice. As comparedwith the control, after spraying the silica sol, selenium sol, andselenium composite silica sol foliage blocking agents, the cadmiumcontents in the early mature rices in 2014 were reduced by 64.3, 49.0and 85.9% respectively, and the cadmium contents in the late maturerices were reduced by 34.2%, 28.8% and 46.4% respectively. Spraying theselenium composite silica sol foliage blocking agent had significantlybetter effects on the reduction of the cadmium content in rice over thesilica sol and selenium sol foliage blocking agents; and spraying thefoliage blocking agent twice during the rice growth phase hadsignificantly better effects on reduction of the cadmium content in riceover solely spraying the same once. In the early mature rice in 2014,after spraying the selenium composite silica sol foliage blocking agentonce for each of the periods from the tillering stage to the jointingstage, and from the heading stage to the filling stage, the cadmiumcontent in rice can be reduced from 1.51 mg/kg in the control to 0.162mg/kg, which had reached the Food Sanitation Standards. Therefore, thefoliage blocking technology for precise regulation of the expression ofthe cadmium absorption and transportation-related gene in rice canachieve the safety of the rice production in the low-to-mediatecontaminated rice field.

TABLE 13 The effects of various foliage blocking materials on thecadmium accumulation levels in rice cadmium content in Cadmium contentin early mature rice late mature rice Reduction as Reduction as Cadmiumcompared Cadmium compared content with control content with control(mg/kg) (%) (mg/kg) (%) Control 1.151 ± 0.074 0.0 1.283 ± 0.098 0.0Silica sol 0.411 ± 0.006 64.3 0.844 ± 0.074 34.2 Selenium 0.587 ± 0.01449.0 0.914 ± 0.076 28.8 sol Selenium 0.162 ± 0.049 85.9 0.688 ± 0.04846.4 composite silica sol

TABLE 14The primers for identification of the cadmium, silicon and arsenictransportation-related genes in rice Gene name Forward primerReverse primer OsNramp5 TGAGCTGCTCTGGGTGATTC TGCCAGCAGCCATAGGAAAA OsLCT1TGGCGATCTTTGGAGGCTTT CGCCGAGGTCGATAAGAACA OsHMA3 AGAACAGCAGGTCGAAGACGATTGCTCAAGGCCATCTGCT OSLsi1 CGGTGGATGTGATCGGAACCA CGTCGAACTTGTTGCTCGCCAOSLsi2 ATCTGGGACTTCATGGCCC ACGTTTGATGCGAGGTTGG

The above examples are the preferred embodiments in the presentinvention, but the embodiments of the present invention are not limitedthereto. Any other changes, modifications, alternatives, combinations,simplifications, made without departing from the spirit and principlesof the present invention, should be the equivalent replacement mode, andincluded in the scope of protection of the present invention.

1. A method for preparing a foliage blocking agent for preciseregulation of the expression of the cadmium absorption andtransportation-related gene in rice, comprising the steps of: (1)preparing a nano-sized selenium gel comprising adding an ascorbic acidsolution into a selenium-containing solution in a water bath at 25-50°C., reacting the ascorbic acid solution and the selenium-containingsolution for 2-5 h, and separating and purifying products, so as toobtain a nano-sized selenium gel; and (2) dispersing the nano-sizedselenium gel comprising adding an emulsifying agent to the nano-sizedselenium gel prepared in step (1) under stirring, and adjusting the pHto 4.5-6.5, wherein the foliage blocking agent for precise regulation ofthe expression of the cadmium absorption and transportation-related genein rice is obtained.
 2. The method of claim 1, wherein theselenium-containing solution in step (1) comprises at least one ofseleninic acid, sodium selenite and potassium selenite.
 3. The method ofclaim 1, wherein a molar ratio between ascorbic acid in the ascorbicacid solution and selenium in the selenium-containing solution in step(1) is (1:1)-(1:3).
 4. The method of claim 1, wherein a finalemulsifying agent content in step (2) is 0.05˜5% by mass; and thefoliage blocking agent in step (2) has a selenium content of 0.15˜2% bymass.
 5. A foliage blocking agent for precise regulation of theexpression of the cadmium absorption and transportation-related gene inrice, characterized in that the foliage blocking agent is prepared bythe method according to claim
 1. 6. A selenium composite silica solfoliage blocking agent for precise regulation of the expression of thecadmium absorption and transportation-related gene in rice, comprisingthe foliage blocking agent in claim 5 and a silica sol.
 7. The seleniumcomposite silica sol foliage blocking agent of claim 6, wherein theselenium composite silica sol foliage blocking agent has a silicacontent of 3˜10% by mass; and a mass ratio between selenium and thesilica in the selenium composite silica sol foliage blocking agent is(1:10)-(1:55).
 8. The selenium composite silica sol foliage blockingagent of claim 6, wherein the silica sol is an acidic silica sol; theacidic silica sol has a pH of 4.5-6.5, a silica content of 15%-20% bymass; and the method for preparing the acidic silica sol, comprises thesteps of: adding a metallic silicon powder or a metal silicate to analkaline solution in a water bath at 40-60° C., at a stirring speed of0.5˜5 m/s, so as to obtain an alkaline silicon-containing solution;passing the alkaline silicon-containing solution through a hydrogen-typeweak acid cation exchange resin column at a speed of 1˜10 mL/min, andcontrolling the pH value of the collection liquid at the column outletto 4.5˜6.5, so as to obtain an active acidic silica sol precursor; andheating the active acidic silica sol precursor to 40˜60° C. in amicrowaver or water bath, maintaining temperature and stirring for 10˜30min, then cooling for 30˜60 min, so as to prepare the acidic silica sol.9. (canceled)
 10. A foliage blocking method for precise regulation ofthe expression of the cadmium absorption and transportation-related genein rice, comprising the steps of: diluting the foliage blocking agentaccording to claim 5, the selenium composite silica sol foliage blockingagent according to claim 6, or a silica sol with water to obtain adiluted foliage blocking agent, diluted selenium composite silica solfoliage blocking agent or diluted silica sol, atomizing the dilutedfoliage blocking agent, diluted selenium composite silica sol foliageblocking agent or diluted silica sol, and spraying the diluted foliageblocking agent, diluted selenium composite silica sol foliage blockingagent or diluted silica sol once on the rice leaf blades during theperiod from the rice tillering stage to the jointing stage, or from theheading to the filling stage, or once for each period, wherein thespraying is carried out in a sunny day or at 4-6 p.m. on a cloudy day;wherein selenium content in the diluted foliage blocking agent iscontrolled at 0.01%˜0.1% by mass; wherein silica content in the dilutedsilica sol or diluted selenium composite silica sol foliage blockingagent is controlled at 0.1%˜0.1% by mass; wherein a spraying dosageduring the period from the tillering stage to the jointing stage is80˜120 liters of the diluted foliage blocking agent, diluted silica solor diluted selenium composite silica sol foliage blocking agent per mu;and wherein a spraying dosage during the period from the heading stageto the filling stage is 100˜150 liters of the diluted foliage blockingagent, diluted silica sol, or diluted selenium composite silica solfoliage blocking agent per mu.